Lipids are known to be useful as carriers for the delivery of drugs to mammals including humans. In pharmaceutical preparations lipids are variously used as admixtures with drugs or in the form of liposomes.
Liposomes are vesicles comprising closed bilayer membranes containing an entrapped aqueous phase. Liposomes may be any variety of unilamellar vesicles (possessing a single membrane bilayer) or multilamellar vesicles (e.g. onion-like structures characterized by concentric membrane bilayers, each separated from the next by an aqueous layer).
Liposomes are formed by methods well known in the art. The original liposome preparation of Bangham et al. (1965, J. Mol. Biol. 13:238-252) involves suspending phospholipids in an organic solvent which is then evaporated to dryness leaving a phospholipid film on the reaction vessel. Then an appropriate amount of aqueous phase is added, the mixture is allowed to "swell", and the resulting liposomes which consist of multilamellar vesicles are dispersed by mechanical means. The structure of the resulting membrane bilayer is such that the hydrophobic (nonpolar) "tails" of the lipid orient toward the center of the bilayer while the hydrophilic (polar) "heads" orient toward the aqueous phase. This technique provides the basis for the development of the small sonicated unilamellar vesicles described by Papahadjopoulos and Miller (1967, Biochim. Biophys. Acta. 135:624-638) and large unilamellar vesicles.
Another class of liposomes is characterized as having substantially equal interlamellar solute distribution. This class of liposomes is denominated as stable plurilamellar vesicles (SPLV) as defined in U.S. Pat. No. 4,522,803 to Lenk et al. and includes monophasic vesicles as described in U.S. Pat. No. 4,588,578 to Fountain et al. and frozen and thawed multilamellar vesicles (FATMLV) as described in "Solute Distributions and Trapping Efficiencies Observed in Freeze-Thawed Multilamellar Vesicles," Mayer et al., Biochima et Biophysica Acta. 817:193-196 (1985).
Another method of liposome formation is by the infusion of lipid solvent such as diethyl ether or ethanol which contains phospholipids into an aqueous solution containing a pharmacological agent resulting in the formation of liposomes which entrap a portion of the aqueous solution. This procedure cannot be used to entrap lipid soluble pharmacological agents soluble in fat or fat solvents due to the very limited solubility of such agents in an aqueous solution.
Lipid soluble pharmacological agents include anti-neoplastics such as doxorubicin; antifungals such as miconazole, terconazole and amphotericin B; immunomodulators such as cyclosporin A; derivatives of muramyl dipeptides such as muramyl tripeptide phosphatidylethanolamine; and, hormones such as glucocorticoids, mineralocorticoids and estrogens; anti-inflammatories such as the steroidals, prednisone, dexamethasone and fluromethasone and the nonsteroidals indomethacin, salicylic acid acetate (aspirin) and ibuprofen, further including analgesic agents such as acemetacin and flurobiprofen; and other agents such as lipoxygenase inhibitors, prostaglandins, neuroleptics, antidepressants, fat-soluble vitamins, contrast materials and antivirals. Pharmacological agents as used herein includes agents administered to animals including mammals, particularly humans, in the course of treatment or diagnosis. Biologically active materials such as drugs as well as diagnostic agents and contrast materials which are usually nonreactive are all to be understood to be pharmacological agents.
Solubilization of lipid soluble pharmacological agent-lipid suspension preparation in water is usually done with the help of solubilizing agents such as polyethylene glycols and propylene glycol, or via surfactants including such well known surfactants as polysorbates, poloxamers, and polyethoxylated castor oil. Upon administration, however, these agents may be present in concentrations sufficient to induce undesirable side effects.
To avoid the use of such agents, D. Schmidt (U.S. Pat. No. 4,271,196) proposed colloidal suspensions formed by solubilization of lipids in ethanol, removal of the solvent by evaporation and addition of water or buffer with the drug added before water or in the colloidal suspension of lipids. Similarly, J. Schrank and H. Steffen (U.S. Pat. No. 4,411,894) solubilized both lipids and drug in ethanol, then ethanol was removed and buffer was added to form liposomes.
These and other procedures involving the removal of ethanol and liposome formation have two major disadvantages. First, ethanol cannot solubilize certain lipids; in particular, salt forms of acidic, or basic phosphatides ("charged phosphatides") such as phosphatidic acid, dicetylphosphate, phosphatidylethanolamine, and phosphatidylserine. Second, the entrapment of lipophilic drug in liposomes is limited such that the drug/lipid ratio (wt/wt) is usually less than 0.2.
It is to be understood that neutral lipids are those which do not present a charge at neutral pH. Phospatidylcholine having a zwitterionic group is termed a neutral polar lipid and compounds such as cholesterol or triglycerides are nonionizable at physiological pH's and are termed neutral nonpolar lipids.
To increase the efficacy of drug solubilization by the lipids, F. Tsunekazu et al. (European Pat. No. 0161445A1) proposed the solubilization of lipids and drug in an organic solvent, removal of the organic solvent, homogenization of the resulting film in aqueous solution by ultrasonic treatment, centrifugation of the suspension and recovery of the lower most layer of the sediment, to yield a particular drug-phospholipid complex. In this publication, particular reference is made to drugs having a molecular weight below 1,000.
Lipid preparations such as liposomes carrying pharmacological agent-lipid solution agents are often characterized by having insufficient shelf life. Dried liposome preparations have been offered to overcome this problem however such preparations must be reconstituted at the time of use. Reconstitution may be associated with problems of clumping and uncertainty as to the liposomal size of the reconstituted preparation, and uncertainty as to the strength of an aliquot. These preparations are also associated with rapid sedimentation.
It is an object of this invention to provide a pharmacological agent-lipid solution preparation in high drug to lipid ratio.
It is a further object of this invention to provide a pharmacological agent-lipid solution preparation wherein the pharmacological agent is of a molecular weight of greater than about 1000.
It is another object of this invention to provide a pharmacological agent-lipid solution preparation sterilizable by filtration.
It is an additional object of this invention to provide a pharmacological agent-lipid solution preparation of lipophilic pharmacological agent.
It is another object of this invention to provide a pharmacological agent-lipid solution preparation that will form a suspension of lipid associated with said pharmacological agent upon introduction into an aqueous medium and further that such suspension exhibit a stability of at about least 0.25 to 6 hours or longer without sedimentation and preferably at least about 2 hours.
It is a further object of this invention to provide a method of forming such suspension.
It is another object of this invention to provide a method of treating mammals, including humans, with therapeutically effective amounts of such suspension.